Oridonin induces apoptosis and reverses drug resistance in cisplatin resistant human gastric cancer cells

Abstract

Gastric cancer is the third most frequent cause of cancer-associated mortality and almost all patients who respond initially to cisplatin (DDP) later develop drug resistance, indicating multi-drug resistance (MDR) is an essential aspect of the failure of treatment. The natural diterpenoid component Oridonin (Ori) has exhibited efficient inhibition in several types of human cancer. However, the effect and potential mechanism of Ori-reversed MDR in human gastric cancer has not been fully elucidated. In the present study, it was found that Ori significantly suppressed DDP-resistant human SGC7901/DDP cell proliferation, growth and colony formation, causing increased caspase-dependent apoptosis, decreased expression of P-glycoprotein (P-gp), encoded by the MDR gene, multi-drug resistance-associated protein (MRP1), and cyclin D1. SGC7901/DDP cells were cultured with different groups of drugs (Ori, DDP alone, or the combination of Ori and DDP). The drug sensitivity, cell apoptosis and effects on MDR were detected by MTT assay and western blot analysis. The results revealed that Ori is able to reverse the DDP resistance and has a clear synergistic effect with DDP in SGC7901/DDP cells by decreasing the levels of P-gp, MRP1, cyclin D1 and cancerous inhibitor of protein phosphatase 2A. Thus, Ori may be a novel effective candidate to treat DDP-resistant human gastric cancer cells.

Keywords: apoptosis; gastric cancer; multi-drug resistance; oridonin.

Figure 1.

Inhibitory effects of Ori on gastric cancer cells. (A) Chemical structure of Ori. (B and C) The inhibitory effects of Ori on SGC7901 and SGC7901/DDP cells, as analyzed by MTT assay. (D) Inhibitory effects of Ori on cell viability of SGC7901/DDP cells assayed by trypan blue exclusion assay. (E) The colony formation assays of SGC7901/DDP cells treated with Ori at indicated concentration. Ori, oridonin.

Figure 2.

The effects of Ori on apoptosis. (A) SGC7901/DDP cells were incubated with various doses of Ori for 24 h. Cells were examined by DAPI staining. Scale bar=200 µm (B) SGC7901/DDP cells were treated with increasing concentrations of Ori for 24 h. Western blot was conducted using antibodies indicated. Ori, oridonin; PARP, poly (ADP-ribose) polymerase; Pro-cas-3, pro-caspase-3.

Figure 3.

The effects of Ori on multidrug resistance proteins. SGC7901/DDP cells were treated with increasing concentrations of Ori for 24 h. Western blot analysis was conducted using the indicated antibodies. Ori, oridonin; MRP1, multi-drug resistance-associated protein; P-gp, P-glycoprotein.

Figure 4.

The effects of Ori on Akt signaling associated molecules. SGC7901/DDP cells were treated with increasing concentrations of Ori for 24 h. Western blot analysis was conducted using the indicated antibodies. Ori, oridonin; CIP2A, cancerous inhibitor of protein phosphatase 2A; PP2Ac, catalytic subunit of protein phosphatase 2A; pAkt, phosphorylated Akt; ERK1/2, extracellular signal-regulated kinase 1/2; pERK1/2, phosphorylated ERK1/2.

Figure 5.

Ori exerts synergistic effects in combination with DDP. (A) SGC7901/DDP cells were treated for 24 h with Ori and/or DDP, and then assessed by MTT assay. *P<0.05; **P<0.01. (B) SGC7901/DDP cells were treated with Ori (O; 20 µM), DDP (D; 20 µM) alone or together (O+D) for 24 h. The treated cells were collected, lysed and assessed by western blot analysis using the indicated antibodies. DDP, cisplatin; Ori oridonin; CIP2A, cancerous inhibitor of protein phosphatase 2A; PP2Ac, catalytic subunit of protein phosphatase 2A; MRP1, multi-drug resistance-associated protein; P-gp, P-glycoprotein; PARP, poly (ADP-ribose) polymerase; Pro-cas-3, pro-caspase-3.